- Email: [email protected]
Paediatric Laboratory Medicine — Some reflections on the sub-specialty
Paediatric Laboratory Medicine- some reflections on the sub-specialty V.L. Grey, T.P. Loh, M. Metz, T. Lang, M. Hersberger PII: DOI: Reference:
S0009-9120(17)30358-2 doi:10.1016/j.clinbiochem.2017.04.005 CLB 9522
To appear in:
Clinical Biochemistry
Accepted date:
10 April 2017
Please cite this article as: Grey VL, Loh TP, Metz M, Lang T, Hersberger M, Paediatric Laboratory Medicine- some reflections on the sub-specialty, Clinical Biochemistry (2017), doi:10.1016/j.clinbiochem.2017.04.005
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
ACCEPTED MANUSCRIPT Paediatric Laboratory Medicine- some reflections on the sub-specialty.
RI
Department of Pathology and Molecular Medicine, McMaster University, Hamilton, ON,
SC
1
PT
V.L.Grey1, T.P.Loh2, M.Metz3, T.Lang4, M.Hersberger5.
Canada, 2Department of Laboratory Medicine, National University Hospital, Singapore, Consultant Chemical Pathologist, SAPath at The Women's & Children's Hospital, North
NU
3
MA
Adelaide, SA, Australia, 4Department of Clinical Biochemistry, University Hospital of North Durham, Durham, DH1 5TW, UK, 5Division of Clinical Chemistry and Biochemistry, University
1
AC CE P
TE
D
Children's Hospital Zurich, Steinwiesstrasse 75, 8032 Zurich, Switzerland
Corresponding author: Dr. VijayLaxmi Grey
McMaster University
Department of Pathology and Molecular Medicine 1280 Main Street West, HSC-2N16 Hamilton ON Canada L8S 4K1
Email: [email protected]
Fax: 905-521-5099
ACCEPTED MANUSCRIPT In many countries there is a tradition of care for children in a children’s hospital. Wikipedia lists 31 countries with Children’s hospitals. In Australia, the combined population
PT
of nearly 30 million people (1) is served by 12 children’s hospitals with each centre of more
RI
than 1 million people having at least one children’s hospital. In Australia children rarely
SC
attend general hospitals. In Canada paediatric tertiary care is provided in children’s hospitals but many community hospitals also offer paediatric care. This is true in several
NU
countries in Europe, USA, and the United Kingdom. Dedicated paediatric services are only
MA
available in tertiary-care hospitals in Singapore. In this environment a strong tradition of paediatric laboratory medicine, providing diagnostic services for babies and children has
TE
AC CE P
laboratory medicine.
D
developed. This article will discuss some key aspects of the practice of paediatric
The goal of laboratory testing is to support disease management. In neonates, infants and children this can present special challenges (Table 1). The first of these is the availability of a suitable sample for testing. When frequent testing is required as may happen when adaptation to extra-uterine life is compromised (as in neonatal hypoglycemia) or a growing child is acutely ill, frequent blood draws may be required. Since less blood is available for testing– a 10mL tube of blood may be as much as 10% of the baby’s total blood volume- judicious test selection is necessary (2). As a general rule no more than 5% of the total infant blood volume should be removed in a single blood draw. The higher hematocrit in neonates also results in less serum volume for testing (2). In general the aim should be to reduce the risk of iatrogenic anaemia due to phlebotomy.
ACCEPTED MANUSCRIPT Both line draws and skin puncture are often used for testing in children. In both cases care should be taken to avoid contamination of the sample: line draws dilution with
PT
heparin or contamination with the infusate will affect the results, and with skin puncture
RI
contamination with interstitial fluid can occur with too much massaging of the site. The use
SC
of EMLA (prilocaine-lidocaine) cream, a topical anaesthetic frequently used in infants and children prior to skin puncture can interfere with analysis of skin biopsies (3) and
NU
acylcarnitines (4). Heel pricks are commonly used to collect blood in neonates and one
MA
should follow the guidelines (5,6) in place for safe skin puncture. Other sample types, such as urine collection can also be problematic. For a random collection, many centres allow
D
collecting the sample on cotton balls or gauze placed inside the diaper. The laboratory
TE
should perform validation of this approach to confirm the acceptability of samples for any
AC CE P
given analysis. In many instances a 24hour collection is needed to provide better diagnostic discrimination but this is challenging even within a hospital setting. Reporting urine analytes in relationship to creatinine concentration is a way to facilitate the use of random samples.
The development of micro-analytical procedures has facilitated small volume testing for routine tests in recent years. Many chemistry analyzers can perform routine tests using only 10 µL of sample. Dead volume may vary between instruments so it is important when investigating a new analyzer to consider both requirements. The availability of robotic systems to handle small volume microtainers is still a challenge for manufacturers. Another important aspect is in the appropriate choice of quality control
ACCEPTED MANUSCRIPT materials. These should take into consideration the critical cut-points relevant to paediatric laboratory testing. In the neonate, bilirubin concentrations are high in the first few days of
PT
life and glucose concentrations can be low. Rapid turnaround time is also important for
RI
some tests in children. This is extremely helpful for clinical decision-making and treatment
SC
strategies especially in the young child when only a limited physical examination and history may be possible. The continued advancements in micro-fluidics and nano-scale
NU
engineering may see more laboratory tests being made available in the near future on
MA
point-of-care (POC) devices that has minimal dead volume and rapid turnaround time.
D
The test menu in paediatric laboratories differs from that of adult laboratories often
TE
because of diseases considered to be primarily that of childhood. Some of these tests are
AC CE P
not frequently ordered and so may not be cost effective when done in the local laboratory. Predominant amongst these are the biochemical tests for the diagnosis of inborn errors of metabolism and follow up of positive newborn screening. Newborn screening has been a public health success in many countries and identifies asymptomatic newborn conditions for early diagnosis and management. It is important to have a good network of reference laboratories for the evaluation of rare diseases. The orphanet initiative (www.orpha.net), allows searching for orphan diseases and directly lists the specialized pediatric laboratories offering the appropriate diagnostic procedures. Groups such as ERNDIM (European Research Network for evaluation and improvement of Screening, Diagnosis and treatment of Inherited Disorders of Metabolism) provide worldwide support for quality assurance in laboratory testing for IEM through their many quality control schemes.
ACCEPTED MANUSCRIPT Newer technologies such as Next Generation Sequencing (NGS) have the promise for diagnosis and discovery of rare genetic diseases. Given that NGS has the potential to
PT
identify untreatable illnesses, and variants of unknown status, best practice guidelines are
RI
required before implementation. Groups such as the UK National Metabolic Biochemistry
SC
Network (MetBioNet) or the Swiss Group for Inborn Errors of Metabolism (SGIEM) support specialised laboratories providing services for the diagnosis and management of Inherited
NU
Metabolic Disease (IMD). Not only do they develop best practice guidelines for the
MA
investigation and management of IMDs, but also they play an important role in training of post registration specialist scientists and establishment of quality assurance programmes.
D
Education is also one of the goals of the Society for the Study of Inborn Errors of
TE
Metabolism (SSIEM) and this is achieved through the publication Journal of Inherited
AC CE P
Metabolic Disease and its annual meetings. Other exciting new areas for the application of NGS in paediatric laboratory medicine include diagnosis, treatment and monitoring of childhood malignancy (precision medicine) (7-9), and NGS meta-genomic sequencing for diagnosing previously hard to detect pathogens (10- 14). The latter may be particularly helpful in the investigation of fever of unknown origin in children and situations where clinical presentation is ambiguous.
Another major challenge is in the interpretation of laboratory tests. This must be done within the context of growth and development. This is an area where laboratory professionals working in paediatric environment can add the most clinical value by actively collaborating with our clinical colleagues in complex/ dynamic testing, result interpretation,
ACCEPTED MANUSCRIPT implementing reflect/ reflex testing and developing appropriate clinical/ testing protocol. Such activities can happen as clinical consultation or multi-disciplinary meetings. An
PT
excellent example of such activity is the development of a protocol for the investigation of
RI
hypoglycaemia in childhood (15). Reliable reference intervals are important for the correct
SC
interpretation of laboratory test results and clinical decision-making. Laboratories should provide age-specific reference intervals, and in the peri-natal period this should also be
NU
gestational age specific. The influence of prematurity on laboratory test results is still not
MA
well understood. The variation in thyroid stimulating hormone is an excellent example of an analyte in which we see changes in cut-points depending on gestational age and time
D
after birth. In recent years several studies such as CALIPER (Canadian Laboratory Initiative
TE
on PEdiatric Reference Intervals)(16), and KIGGS (the German survey on children’s health
AC CE P
as database for reference intervals) (17) have tried to address this problem but there are still significant gaps in the knowledge. In Australia, increased effort in the development of reference intervals has occurred recently with both a priori (18) and post priori (19) schemes employed. Harmonised reference intervals for children for ten common analytes have been developed and disseminated through the paediatric special interest group (PSIG) of the AACB (Australasian Association of Clinical Biochemists).
Population reference intervals remain an ongoing challenge even in adults. It has been shown that there are fundamental differences among adult Asian populations, which preclude the use of harmonised reference intervals for certain biochemistry tests (20). However, for countries with more homogeneous population like Japan, the use of
ACCEPTED MANUSCRIPT harmonised reference intervals is possible (21). Direct derivation of paediatric reference intervals is relatively under-developed in Asia. Most of the paediatric reference intervals
PT
developed in this region tend to be sporadic, and initiated by a single centre. They appear
RI
to address a specialised local clinical need instead of focusing on general biochemistry.
SC
More recently, there are some efforts to use indirect (data mining) methods to derive paediatric reference intervals (19,22) and biological variation data (22-24).
NU
Notwithstanding their limitations, these relatively more accessible methods can pave the
MA
way for laboratories in resource-restricted regions to verify, develop and validate paediatric reference intervals. Children change everyday and laboratory reference intervals
D
should reflect that. This can be achieved by expressing reference values as smoothed
TE
continuous variable with age in a manner similar to growth charts for children. Laboratory
AC CE P
information system serving paediatric patients should facilitate this practice to improve the clinical accuracy of the age-specific reference values. Examples of where this can be helpful are the management of neonatal hyperbilirubinemia when hour–specific values are important for the assessment of risk, and other analytes such as alkaline phosphatase and sex hormones that vary throughout childhood and puberty.
Cut-points in paediatric diseases are also difficult to find. Consensus guidelines are established for adults but information on children is still limited. While several risk scores are available to assess the global cardiovascular risk for adults, there are none based on cardiovascular morbidity and mortality from prospective studies in children. Exceptions
ACCEPTED MANUSCRIPT are the guidelines for the diagnosis and the management of familial hypercholesterolemia
PT
in children, which have gone from no guidelines ten years ago to several now (25,26).
RI
Critical values also differ in a paediatric population because of the less robust
SC
homeostasis and lesser reserves in the neonate and infant. In a recent survey by the IFCC Taskforce on Paediatric Laboratory Medicine, more than 70% of the laboratories reported
NU
critical values for sodium, potassium, glucose, and calcium but there was a lack of
MA
agreement on the values reported. In addition, the number of analytes included in the list for critical values and the range of cut-off values applied vary widely. In part, this is due to
D
the lack of paediatric critical values that are defined by robust evidence based on clinical
AC CE P
TE
outcomes (27). Clearly this is another area that requires collaboration.
The practice of Paediatric Laboratory Medicine offers many opportunities for collaborations in research, clinical practice and education. Paediatric special interest groups within many national societies such as the Pediatric Maternal Fetal division of the American Association of Clinical Chemistry (1983), the pediatric focus group of the Canadian Society of Clinical Chemistry (1994), and more recently the paediatric special interest group (PSIG) of the AACB (2012), the Association of Clinical Biochemistry and Laboratory Medicine (ACB) established a Paediatric Laboratory Medicine Network (2014) and the SGIEM (2015) foster these collaborations. In 1980, a small group of individuals with experience in the field held the first of a series of congresses of the International Congress of Paediatric Laboratory Medicine in Jerusalem. Since this time meetings have been held
ACCEPTED MANUSCRIPT every 2-3 years in different cities in the world. In 1995, the International Association of Paediatric Laboratory Medicine was inaugurated (28). The association came under the
PT
umbrella of the IFCC in 2006 and is now the IFCC Task Force of Paediatric Laboratory
RI
Medicine (TF-PLM). The members of the Task Force are representative of paediatric
SC
laboratory specialists from different national societies. One of the aims of the Task Force is to create a worldwide network of scientists working in laboratories specialized in Pediatric
MA
NU
Medicine.
In a survey of IFCC member societies done by the TF-PLM in 2012, the majority of
D
respondents wanted to learn more about pediatric laboratory medicine. The International
TE
Congress of Pediatric Laboratory Medicine organized by the TF-PLM and held every three
AC CE P
years in different countries just prior to the IFCC WorldLab is one venue that provides up to date information. Further avenues such as a curriculum for trainees in the field of laboratory medicine for disseminating education in the field are currently being developed. The need for education in the field cannot be over-emphasized. While in the developed world diagnostic services for children are more readily available, it is not easily accessed in many countries. Activities of the Task Force also include efforts to coordinate activities worldwide towards the establishment of reference intervals for laboratory test results in pediatric patients of all age groups, and an approach to the development of pediatric critical values.
ACCEPTED MANUSCRIPT Paediatric Laboratory Medicine is entering an exciting era. Breakthrough advances in bioengineering (microfluidics, nano-manufacturing), information technology (big data
PT
analysis), genetic testing (NGS) and gene editing (CRISPR) promise to overcome many
RI
challenges that seemed insurmountable previously. Moreover, many longitudinal studies
SC
that incorporate genetic, environment and lifestyle factors are being conducted worldwide to learn about how they affect the growth and development of children. Together, these
NU
advancements will bring unprecedented insights into the health and disease of childhood.
MA
The global trend of declining fertility rate has meant that more attention and resources are focused on the smaller number of precious children. The role of the Paediatric Laboratory
D
Medicine community is to keep abreast to these developments and facilitate the
TE
translation of these advances into clinical practice by active collaboration with our clinical
AC CE P
and laboratory colleagues.
ACCEPTED MANUSCRIPT References
PT
1. Australian Bureau of Statistics accessed 28.11.2016 on www.abs.gov.au/Population
RI
2. Green A, Morgan I, Gray J, Neonatal and Laboratory Medicine, eds, Harris B, Marshall W,
SC
2003; Chapter 1,11-12
3. Vallance H, Chaba T, Clarke L, Taylor G. Pseudo-lysosomal storage disease caused by
NU
EMLA cream, J.Inherit Metab Dis 2004;27, 507-11
MA
4. Kuster T, Torresani T, Kleinert P, et al Filter paper cards contaminated with EMLA cream produce artefacts on acylcarnitine analysis, J.Inherit Metab Dis 2004;27:707-9
D
5. Procedures and Devices for the Collection of Diagnostic Capillary Blood specimens;
TE
approved standard- 6th edition, CLSI guideline H04-A6; Vol. 28 No25
AC CE P
6. WHO Guidelines on Drawing Blood: Best Practices in Phlebotomy Geneva: World Health Organization; 2010- Paediatric and neonatal blood sampling p51- 63 7. van Wezel EM, Zwijnenburg D, Zappeij-Kannegieter L, et al . Whole-genome sequencing identifies patient-specific DNA minimal residual disease markers in neuroblastoma. J Mol. Diagn. 2015 Jan; 17(1): 43-52
8. Oberg JA, Glade Bender JL, Sulis ML, et al. Implementation of next generation sequencing into pediatric hematology-oncology practice: moving beyond actionable alterations. Genome Med. 2016 Dec 23;8(1): 133. 9. Kline CN, Joseph NM, Grenert JP, et al. Targeted next-generation sequencing of pediatric neuro-oncology patients improves diagnosis, identifies pathogenic germline mutations, and directs targeted therapy. Neuro Oncol. 2016 Nov 14. pii: now254. [Epub ahead of print]
ACCEPTED MANUSCRIPT 10. Graf EH, Simmon KE, Tardif KD, et al. Unbiased Detection of Respiratory Viruses by Use of RNA Sequencing-Based Metagenomics: a Systematic Comparison to a Commercial PCR
PT
Panel. J Clin Microbiol. 2016 Apr;54(4):1000-7.
RI
11. Fukui Y, Aoki K, Okuma S, Sato T, Ishii Y, Tateda K. Metagenomic analysis for detecting
SC
pathogens in culture-negative infective endocarditis. J Infect Chemother. 2015 Dec; 21(12): 882-4.
NU
12. Feigelman R, Kahlert CR, Baty F, et al Sputum DNA sequencing in cystic fibrosis: non-
MA
invasive access to the lung microbiome and to pathogen details. Microbiome. 2017 Feb 10; 5(1):20.
D
13. Doan T, Wilson MR, Crawford ED, et al Illuminating uveitis: metagenomic deep
TE
sequencing identifies common and rare pathogens. Genome Med. 2016 Aug 25;8(1):90.
AC CE P
14. Ward DV, Scholz M, Zolfo M, et al Metagenomic Sequencing with Strain-Level Resolution Implicates Uropathogenic E. coli in Necrotizing Enterocolitis and Mortality in Preterm Infants. Cell Rep. 2016 Mar 29;14(12):2912-24. 15. Lang TF. Update on investigating hypoglycaemia in childhood. Ann Clin Biochem. 2011 May; 48(Pt 3): 200-11.
16. Colantonio D, Kyriakopoulou L, Chan MK et al. Closing the gaps in Paediatric laboratory reference intervals: a CALIPER database of 40 biochemical markers in a healthy and multiethnic population of children. Clin Chem 2012;58: 854-68. 17. Kohse K, Thamm M. KIGGS-the German survey on children’s health as database for reference intervals. Clin Biochem 2011; 44:479
ACCEPTED MANUSCRIPT 18. Southcott EK, Kerrigan JL, Potter JM et al. Establishment of pediatric reference intervals on a large cohort of healthy children. Clin Chim Acta 2010 411: 1421-7
PT
19. Loh TP, Antoniou G, Baghurst P, Metz MP. Development of paediatric biochemistry
RI
centile charts as a complement to laboratory reference intervals. Pathology. 2014
SC
Jun;46(4):336-43.
20. Ichihara K, Itoh Y, Lam CWK, et al.: Science Committee for the Asian-Pacific Federation
NU
of Clinical Biochemistry. Sources of variation of commonly measured serum analytes in 6
MA
Asian cities and consideration of common reference intervals. Clin Chem 2008; 54:356-365. 21. Yamamoto Y, Hosogaya S, Osawa S, Ichihara K, Onuma T, Saito A, et al. Nationwide
D
multicenter study aimed at the establishment of common reference intervals for
TE
standardized clinical laboratory tests in Japan. Clin Chem LabMed 2015;51:1663-1672.
AC CE P
22. Yang Q, Lew HY, Peh RH, Metz MP, Loh TP. An automated and objective method for age partitioning of reference intervals based on continuous centile curves. Pathology. 2016 Oct; 48(6):581-5
23. Loh TP, Metz MP. Indirect estimation of pediatric between-individual biological variation data for 22 common serum biochemistries. Am J Clin Pathol. 2015 May; 143(5):683-93. 24. Loh TP, Ranieri E, Metz MP. Derivation of pediatric within-individual biological variation by indirect sampling method: an LMS approach. Am J Clin Pathol. 2014 Nov;142(5):657-63. 25. Wiegman A, Gidding SS, Watts GF, Chapman MJ, Ginsberg HN, CuchelM, et al European Atherosclerosis Society Consensus Panel, Familial hypercholesterolemia in children and
ACCEPTED MANUSCRIPT adolescents: gaining decades of life by optimizing detection ad treatment. Eur Heart J.2015; 36:2425-37.
PT
26. Watts GF, Gidding S, Wierzbicki AS, Toth PP, Alonso R, Brown WV, Bruckert E, et al,
RI
International Familial Hypercholesterolemia Foundation, Integrated guidance on the care
SC
of familial hypercholesterolemia from the International FH Foundation. Eur J. Prev Cardiol 2015; 22:849-54
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27. Jenkins JJ, Mac Crawford J, Bissell MG. Studying critical values: adverse event
MA
identification following a critical laboratory values study at the Ohio State University Medical Center. Am J Clin Pathol 2007; 128:604-9
D
28. Kohse KP, A worldwide paediatric laboratory medicine network: The International
AC CE P
2009; 62:193-194
TE
Federation of Clinical Chemistry Task Force on Pediatric Laboratory Medicine. J.Clin Pathol
ACCEPTED MANUSCRIPT Table 1 Challenges in the practice of Paediatric Laboratory Medicine
PT
Sample volume must be smallContinue to advocate for instruments to not only support small volume testing but also the handling of small tubes, and
RI
bar codes.
SC
Reference testing for esotericMake information for test referrals available. testing
Continue to monitor for quality assurance of the tests.
NU
Develop guidelines for their use and standardization. Reporting of age-specific values.
Critical values
Promote collaborations for their development of and make
MA
Reference intervals and
them available.
Use mathematical approaches for the development of
D
reference intervals from databases.
Future Developments
TE
Develop standards for reporting age-specific critical values. Collaborate with other pediatric subspecialties to develop
AC CE P
best practice guidelines, and quality assurance protocols for appropriate use of newer technologies in the clinical laboratory.
Education
Make educational materials available on line. Incorporate training for residents and fellows in the field. Continue regular meetings of the ICPLM. Incorporate a symposium on pediatric lab medicine in all WorldLab and National congresses.
S0009-9120(17)30358-2 doi:10.1016/j.clinbiochem.2017.04.005 CLB 9522
To appear in:
Clinical Biochemistry
Accepted date:
10 April 2017
Please cite this article as: Grey VL, Loh TP, Metz M, Lang T, Hersberger M, Paediatric Laboratory Medicine- some reflections on the sub-specialty, Clinical Biochemistry (2017), doi:10.1016/j.clinbiochem.2017.04.005
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
ACCEPTED MANUSCRIPT Paediatric Laboratory Medicine- some reflections on the sub-specialty.
RI
Department of Pathology and Molecular Medicine, McMaster University, Hamilton, ON,
SC
1
PT
V.L.Grey1, T.P.Loh2, M.Metz3, T.Lang4, M.Hersberger5.
Canada, 2Department of Laboratory Medicine, National University Hospital, Singapore, Consultant Chemical Pathologist, SAPath at The Women's & Children's Hospital, North
NU
3
MA
Adelaide, SA, Australia, 4Department of Clinical Biochemistry, University Hospital of North Durham, Durham, DH1 5TW, UK, 5Division of Clinical Chemistry and Biochemistry, University
1
AC CE P
TE
D
Children's Hospital Zurich, Steinwiesstrasse 75, 8032 Zurich, Switzerland
Corresponding author: Dr. VijayLaxmi Grey
McMaster University
Department of Pathology and Molecular Medicine 1280 Main Street West, HSC-2N16 Hamilton ON Canada L8S 4K1
Email: [email protected]
Fax: 905-521-5099
ACCEPTED MANUSCRIPT In many countries there is a tradition of care for children in a children’s hospital. Wikipedia lists 31 countries with Children’s hospitals. In Australia, the combined population
PT
of nearly 30 million people (1) is served by 12 children’s hospitals with each centre of more
RI
than 1 million people having at least one children’s hospital. In Australia children rarely
SC
attend general hospitals. In Canada paediatric tertiary care is provided in children’s hospitals but many community hospitals also offer paediatric care. This is true in several
NU
countries in Europe, USA, and the United Kingdom. Dedicated paediatric services are only
MA
available in tertiary-care hospitals in Singapore. In this environment a strong tradition of paediatric laboratory medicine, providing diagnostic services for babies and children has
TE
AC CE P
laboratory medicine.
D
developed. This article will discuss some key aspects of the practice of paediatric
The goal of laboratory testing is to support disease management. In neonates, infants and children this can present special challenges (Table 1). The first of these is the availability of a suitable sample for testing. When frequent testing is required as may happen when adaptation to extra-uterine life is compromised (as in neonatal hypoglycemia) or a growing child is acutely ill, frequent blood draws may be required. Since less blood is available for testing– a 10mL tube of blood may be as much as 10% of the baby’s total blood volume- judicious test selection is necessary (2). As a general rule no more than 5% of the total infant blood volume should be removed in a single blood draw. The higher hematocrit in neonates also results in less serum volume for testing (2). In general the aim should be to reduce the risk of iatrogenic anaemia due to phlebotomy.
ACCEPTED MANUSCRIPT Both line draws and skin puncture are often used for testing in children. In both cases care should be taken to avoid contamination of the sample: line draws dilution with
PT
heparin or contamination with the infusate will affect the results, and with skin puncture
RI
contamination with interstitial fluid can occur with too much massaging of the site. The use
SC
of EMLA (prilocaine-lidocaine) cream, a topical anaesthetic frequently used in infants and children prior to skin puncture can interfere with analysis of skin biopsies (3) and
NU
acylcarnitines (4). Heel pricks are commonly used to collect blood in neonates and one
MA
should follow the guidelines (5,6) in place for safe skin puncture. Other sample types, such as urine collection can also be problematic. For a random collection, many centres allow
D
collecting the sample on cotton balls or gauze placed inside the diaper. The laboratory
TE
should perform validation of this approach to confirm the acceptability of samples for any
AC CE P
given analysis. In many instances a 24hour collection is needed to provide better diagnostic discrimination but this is challenging even within a hospital setting. Reporting urine analytes in relationship to creatinine concentration is a way to facilitate the use of random samples.
The development of micro-analytical procedures has facilitated small volume testing for routine tests in recent years. Many chemistry analyzers can perform routine tests using only 10 µL of sample. Dead volume may vary between instruments so it is important when investigating a new analyzer to consider both requirements. The availability of robotic systems to handle small volume microtainers is still a challenge for manufacturers. Another important aspect is in the appropriate choice of quality control
ACCEPTED MANUSCRIPT materials. These should take into consideration the critical cut-points relevant to paediatric laboratory testing. In the neonate, bilirubin concentrations are high in the first few days of
PT
life and glucose concentrations can be low. Rapid turnaround time is also important for
RI
some tests in children. This is extremely helpful for clinical decision-making and treatment
SC
strategies especially in the young child when only a limited physical examination and history may be possible. The continued advancements in micro-fluidics and nano-scale
NU
engineering may see more laboratory tests being made available in the near future on
MA
point-of-care (POC) devices that has minimal dead volume and rapid turnaround time.
D
The test menu in paediatric laboratories differs from that of adult laboratories often
TE
because of diseases considered to be primarily that of childhood. Some of these tests are
AC CE P
not frequently ordered and so may not be cost effective when done in the local laboratory. Predominant amongst these are the biochemical tests for the diagnosis of inborn errors of metabolism and follow up of positive newborn screening. Newborn screening has been a public health success in many countries and identifies asymptomatic newborn conditions for early diagnosis and management. It is important to have a good network of reference laboratories for the evaluation of rare diseases. The orphanet initiative (www.orpha.net), allows searching for orphan diseases and directly lists the specialized pediatric laboratories offering the appropriate diagnostic procedures. Groups such as ERNDIM (European Research Network for evaluation and improvement of Screening, Diagnosis and treatment of Inherited Disorders of Metabolism) provide worldwide support for quality assurance in laboratory testing for IEM through their many quality control schemes.
ACCEPTED MANUSCRIPT Newer technologies such as Next Generation Sequencing (NGS) have the promise for diagnosis and discovery of rare genetic diseases. Given that NGS has the potential to
PT
identify untreatable illnesses, and variants of unknown status, best practice guidelines are
RI
required before implementation. Groups such as the UK National Metabolic Biochemistry
SC
Network (MetBioNet) or the Swiss Group for Inborn Errors of Metabolism (SGIEM) support specialised laboratories providing services for the diagnosis and management of Inherited
NU
Metabolic Disease (IMD). Not only do they develop best practice guidelines for the
MA
investigation and management of IMDs, but also they play an important role in training of post registration specialist scientists and establishment of quality assurance programmes.
D
Education is also one of the goals of the Society for the Study of Inborn Errors of
TE
Metabolism (SSIEM) and this is achieved through the publication Journal of Inherited
AC CE P
Metabolic Disease and its annual meetings. Other exciting new areas for the application of NGS in paediatric laboratory medicine include diagnosis, treatment and monitoring of childhood malignancy (precision medicine) (7-9), and NGS meta-genomic sequencing for diagnosing previously hard to detect pathogens (10- 14). The latter may be particularly helpful in the investigation of fever of unknown origin in children and situations where clinical presentation is ambiguous.
Another major challenge is in the interpretation of laboratory tests. This must be done within the context of growth and development. This is an area where laboratory professionals working in paediatric environment can add the most clinical value by actively collaborating with our clinical colleagues in complex/ dynamic testing, result interpretation,
ACCEPTED MANUSCRIPT implementing reflect/ reflex testing and developing appropriate clinical/ testing protocol. Such activities can happen as clinical consultation or multi-disciplinary meetings. An
PT
excellent example of such activity is the development of a protocol for the investigation of
RI
hypoglycaemia in childhood (15). Reliable reference intervals are important for the correct
SC
interpretation of laboratory test results and clinical decision-making. Laboratories should provide age-specific reference intervals, and in the peri-natal period this should also be
NU
gestational age specific. The influence of prematurity on laboratory test results is still not
MA
well understood. The variation in thyroid stimulating hormone is an excellent example of an analyte in which we see changes in cut-points depending on gestational age and time
D
after birth. In recent years several studies such as CALIPER (Canadian Laboratory Initiative
TE
on PEdiatric Reference Intervals)(16), and KIGGS (the German survey on children’s health
AC CE P
as database for reference intervals) (17) have tried to address this problem but there are still significant gaps in the knowledge. In Australia, increased effort in the development of reference intervals has occurred recently with both a priori (18) and post priori (19) schemes employed. Harmonised reference intervals for children for ten common analytes have been developed and disseminated through the paediatric special interest group (PSIG) of the AACB (Australasian Association of Clinical Biochemists).
Population reference intervals remain an ongoing challenge even in adults. It has been shown that there are fundamental differences among adult Asian populations, which preclude the use of harmonised reference intervals for certain biochemistry tests (20). However, for countries with more homogeneous population like Japan, the use of
ACCEPTED MANUSCRIPT harmonised reference intervals is possible (21). Direct derivation of paediatric reference intervals is relatively under-developed in Asia. Most of the paediatric reference intervals
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developed in this region tend to be sporadic, and initiated by a single centre. They appear
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to address a specialised local clinical need instead of focusing on general biochemistry.
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More recently, there are some efforts to use indirect (data mining) methods to derive paediatric reference intervals (19,22) and biological variation data (22-24).
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Notwithstanding their limitations, these relatively more accessible methods can pave the
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way for laboratories in resource-restricted regions to verify, develop and validate paediatric reference intervals. Children change everyday and laboratory reference intervals
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should reflect that. This can be achieved by expressing reference values as smoothed
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continuous variable with age in a manner similar to growth charts for children. Laboratory
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information system serving paediatric patients should facilitate this practice to improve the clinical accuracy of the age-specific reference values. Examples of where this can be helpful are the management of neonatal hyperbilirubinemia when hour–specific values are important for the assessment of risk, and other analytes such as alkaline phosphatase and sex hormones that vary throughout childhood and puberty.
Cut-points in paediatric diseases are also difficult to find. Consensus guidelines are established for adults but information on children is still limited. While several risk scores are available to assess the global cardiovascular risk for adults, there are none based on cardiovascular morbidity and mortality from prospective studies in children. Exceptions
ACCEPTED MANUSCRIPT are the guidelines for the diagnosis and the management of familial hypercholesterolemia
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in children, which have gone from no guidelines ten years ago to several now (25,26).
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Critical values also differ in a paediatric population because of the less robust
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homeostasis and lesser reserves in the neonate and infant. In a recent survey by the IFCC Taskforce on Paediatric Laboratory Medicine, more than 70% of the laboratories reported
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critical values for sodium, potassium, glucose, and calcium but there was a lack of
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agreement on the values reported. In addition, the number of analytes included in the list for critical values and the range of cut-off values applied vary widely. In part, this is due to
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the lack of paediatric critical values that are defined by robust evidence based on clinical
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outcomes (27). Clearly this is another area that requires collaboration.
The practice of Paediatric Laboratory Medicine offers many opportunities for collaborations in research, clinical practice and education. Paediatric special interest groups within many national societies such as the Pediatric Maternal Fetal division of the American Association of Clinical Chemistry (1983), the pediatric focus group of the Canadian Society of Clinical Chemistry (1994), and more recently the paediatric special interest group (PSIG) of the AACB (2012), the Association of Clinical Biochemistry and Laboratory Medicine (ACB) established a Paediatric Laboratory Medicine Network (2014) and the SGIEM (2015) foster these collaborations. In 1980, a small group of individuals with experience in the field held the first of a series of congresses of the International Congress of Paediatric Laboratory Medicine in Jerusalem. Since this time meetings have been held
ACCEPTED MANUSCRIPT every 2-3 years in different cities in the world. In 1995, the International Association of Paediatric Laboratory Medicine was inaugurated (28). The association came under the
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umbrella of the IFCC in 2006 and is now the IFCC Task Force of Paediatric Laboratory
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Medicine (TF-PLM). The members of the Task Force are representative of paediatric
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laboratory specialists from different national societies. One of the aims of the Task Force is to create a worldwide network of scientists working in laboratories specialized in Pediatric
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Medicine.
In a survey of IFCC member societies done by the TF-PLM in 2012, the majority of
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respondents wanted to learn more about pediatric laboratory medicine. The International
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Congress of Pediatric Laboratory Medicine organized by the TF-PLM and held every three
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years in different countries just prior to the IFCC WorldLab is one venue that provides up to date information. Further avenues such as a curriculum for trainees in the field of laboratory medicine for disseminating education in the field are currently being developed. The need for education in the field cannot be over-emphasized. While in the developed world diagnostic services for children are more readily available, it is not easily accessed in many countries. Activities of the Task Force also include efforts to coordinate activities worldwide towards the establishment of reference intervals for laboratory test results in pediatric patients of all age groups, and an approach to the development of pediatric critical values.
ACCEPTED MANUSCRIPT Paediatric Laboratory Medicine is entering an exciting era. Breakthrough advances in bioengineering (microfluidics, nano-manufacturing), information technology (big data
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analysis), genetic testing (NGS) and gene editing (CRISPR) promise to overcome many
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challenges that seemed insurmountable previously. Moreover, many longitudinal studies
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that incorporate genetic, environment and lifestyle factors are being conducted worldwide to learn about how they affect the growth and development of children. Together, these
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advancements will bring unprecedented insights into the health and disease of childhood.
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The global trend of declining fertility rate has meant that more attention and resources are focused on the smaller number of precious children. The role of the Paediatric Laboratory
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Medicine community is to keep abreast to these developments and facilitate the
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translation of these advances into clinical practice by active collaboration with our clinical
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and laboratory colleagues.
ACCEPTED MANUSCRIPT References
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1. Australian Bureau of Statistics accessed 28.11.2016 on www.abs.gov.au/Population
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2. Green A, Morgan I, Gray J, Neonatal and Laboratory Medicine, eds, Harris B, Marshall W,
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2003; Chapter 1,11-12
3. Vallance H, Chaba T, Clarke L, Taylor G. Pseudo-lysosomal storage disease caused by
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EMLA cream, J.Inherit Metab Dis 2004;27, 507-11
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4. Kuster T, Torresani T, Kleinert P, et al Filter paper cards contaminated with EMLA cream produce artefacts on acylcarnitine analysis, J.Inherit Metab Dis 2004;27:707-9
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5. Procedures and Devices for the Collection of Diagnostic Capillary Blood specimens;
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approved standard- 6th edition, CLSI guideline H04-A6; Vol. 28 No25
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6. WHO Guidelines on Drawing Blood: Best Practices in Phlebotomy Geneva: World Health Organization; 2010- Paediatric and neonatal blood sampling p51- 63 7. van Wezel EM, Zwijnenburg D, Zappeij-Kannegieter L, et al . Whole-genome sequencing identifies patient-specific DNA minimal residual disease markers in neuroblastoma. J Mol. Diagn. 2015 Jan; 17(1): 43-52
8. Oberg JA, Glade Bender JL, Sulis ML, et al. Implementation of next generation sequencing into pediatric hematology-oncology practice: moving beyond actionable alterations. Genome Med. 2016 Dec 23;8(1): 133. 9. Kline CN, Joseph NM, Grenert JP, et al. Targeted next-generation sequencing of pediatric neuro-oncology patients improves diagnosis, identifies pathogenic germline mutations, and directs targeted therapy. Neuro Oncol. 2016 Nov 14. pii: now254. [Epub ahead of print]
ACCEPTED MANUSCRIPT 10. Graf EH, Simmon KE, Tardif KD, et al. Unbiased Detection of Respiratory Viruses by Use of RNA Sequencing-Based Metagenomics: a Systematic Comparison to a Commercial PCR
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Panel. J Clin Microbiol. 2016 Apr;54(4):1000-7.
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11. Fukui Y, Aoki K, Okuma S, Sato T, Ishii Y, Tateda K. Metagenomic analysis for detecting
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pathogens in culture-negative infective endocarditis. J Infect Chemother. 2015 Dec; 21(12): 882-4.
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12. Feigelman R, Kahlert CR, Baty F, et al Sputum DNA sequencing in cystic fibrosis: non-
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invasive access to the lung microbiome and to pathogen details. Microbiome. 2017 Feb 10; 5(1):20.
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13. Doan T, Wilson MR, Crawford ED, et al Illuminating uveitis: metagenomic deep
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sequencing identifies common and rare pathogens. Genome Med. 2016 Aug 25;8(1):90.
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14. Ward DV, Scholz M, Zolfo M, et al Metagenomic Sequencing with Strain-Level Resolution Implicates Uropathogenic E. coli in Necrotizing Enterocolitis and Mortality in Preterm Infants. Cell Rep. 2016 Mar 29;14(12):2912-24. 15. Lang TF. Update on investigating hypoglycaemia in childhood. Ann Clin Biochem. 2011 May; 48(Pt 3): 200-11.
16. Colantonio D, Kyriakopoulou L, Chan MK et al. Closing the gaps in Paediatric laboratory reference intervals: a CALIPER database of 40 biochemical markers in a healthy and multiethnic population of children. Clin Chem 2012;58: 854-68. 17. Kohse K, Thamm M. KIGGS-the German survey on children’s health as database for reference intervals. Clin Biochem 2011; 44:479
ACCEPTED MANUSCRIPT 18. Southcott EK, Kerrigan JL, Potter JM et al. Establishment of pediatric reference intervals on a large cohort of healthy children. Clin Chim Acta 2010 411: 1421-7
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19. Loh TP, Antoniou G, Baghurst P, Metz MP. Development of paediatric biochemistry
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centile charts as a complement to laboratory reference intervals. Pathology. 2014
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Jun;46(4):336-43.
20. Ichihara K, Itoh Y, Lam CWK, et al.: Science Committee for the Asian-Pacific Federation
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of Clinical Biochemistry. Sources of variation of commonly measured serum analytes in 6
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Asian cities and consideration of common reference intervals. Clin Chem 2008; 54:356-365. 21. Yamamoto Y, Hosogaya S, Osawa S, Ichihara K, Onuma T, Saito A, et al. Nationwide
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multicenter study aimed at the establishment of common reference intervals for
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standardized clinical laboratory tests in Japan. Clin Chem LabMed 2015;51:1663-1672.
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22. Yang Q, Lew HY, Peh RH, Metz MP, Loh TP. An automated and objective method for age partitioning of reference intervals based on continuous centile curves. Pathology. 2016 Oct; 48(6):581-5
23. Loh TP, Metz MP. Indirect estimation of pediatric between-individual biological variation data for 22 common serum biochemistries. Am J Clin Pathol. 2015 May; 143(5):683-93. 24. Loh TP, Ranieri E, Metz MP. Derivation of pediatric within-individual biological variation by indirect sampling method: an LMS approach. Am J Clin Pathol. 2014 Nov;142(5):657-63. 25. Wiegman A, Gidding SS, Watts GF, Chapman MJ, Ginsberg HN, CuchelM, et al European Atherosclerosis Society Consensus Panel, Familial hypercholesterolemia in children and
ACCEPTED MANUSCRIPT adolescents: gaining decades of life by optimizing detection ad treatment. Eur Heart J.2015; 36:2425-37.
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26. Watts GF, Gidding S, Wierzbicki AS, Toth PP, Alonso R, Brown WV, Bruckert E, et al,
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International Familial Hypercholesterolemia Foundation, Integrated guidance on the care
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of familial hypercholesterolemia from the International FH Foundation. Eur J. Prev Cardiol 2015; 22:849-54
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27. Jenkins JJ, Mac Crawford J, Bissell MG. Studying critical values: adverse event
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identification following a critical laboratory values study at the Ohio State University Medical Center. Am J Clin Pathol 2007; 128:604-9
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28. Kohse KP, A worldwide paediatric laboratory medicine network: The International
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2009; 62:193-194
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Federation of Clinical Chemistry Task Force on Pediatric Laboratory Medicine. J.Clin Pathol
ACCEPTED MANUSCRIPT Table 1 Challenges in the practice of Paediatric Laboratory Medicine
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Sample volume must be smallContinue to advocate for instruments to not only support small volume testing but also the handling of small tubes, and
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bar codes.
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Reference testing for esotericMake information for test referrals available. testing
Continue to monitor for quality assurance of the tests.
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Develop guidelines for their use and standardization. Reporting of age-specific values.
Critical values
Promote collaborations for their development of and make
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Reference intervals and
them available.
Use mathematical approaches for the development of
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reference intervals from databases.
Future Developments
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Develop standards for reporting age-specific critical values. Collaborate with other pediatric subspecialties to develop
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best practice guidelines, and quality assurance protocols for appropriate use of newer technologies in the clinical laboratory.
Education
Make educational materials available on line. Incorporate training for residents and fellows in the field. Continue regular meetings of the ICPLM. Incorporate a symposium on pediatric lab medicine in all WorldLab and National congresses.